Clone, expression and plasminogen binding property of three fructose-1,6-bisphosphate aldolases from Clonorchis sinensis

@# Fructose-1,6-bisphosphate aldolase (FbA), a well characterized glycometabolism enzyme, has been found to participate in other important processes besides the classic catalysis. To understand the important functions of three fructose-1,6-bisphosphate aldolases from Clonorchis sinensis (CsFbAs, CsFbA-1/2/3) in host-parasite interplay, the open reading frames of CsFbAs were cloned into pET30a (+) vector and the resulting recombinant plasmids were transformed into Escherichia coli BL21 (DE3) for expression of the proteins. Purified recombinant CsFbAs proteins (rCsFbAs) were approximately 45.0 kDa on 12% SDS-PAGE and could be probed with each rat anti-rCsFbAs sera by western blotting analysis. ELISA and ligand blot overlay indicated that rCsFbAs of 45.0 kDa as well as native CsFbAs of 39.5 kDa from total worm extracts and excretory-secretory products of Clonorchis sinensis (CsESPs) could bind to human plasminogen, and the binding could be efficiently inhibited by lysine analog ε-aminocaproic acid. Our results suggested that as both the components of CsESPs and the plasminogen binding proteins, three CsFbAs might be involved in preventing the formation of the blood clot so that Clonorchis sinensis could acquire enough nutrients from host tissue for their successful survival and colonization in the host. Our work will provide us with new information about the biological function of three CsFbAs and their roles in hostparasite interplay

Effects of herbal medicine Sijunzi decoction on rabbits after relieving intestinal obstruction

Intestinal obstruction leads to blockage of the movement of intestinal contents. After relieving the obstruction, patients might still suffer with compromised immune function and nutritional deficiency. This study aimed to evaluate the effects of Sijunzi decoction on restoring the immune function and nutritional status after relieving the obstruction. Experimental rabbits (2.5±0.2 kg) were randomly divided into normal control group, 2-day intestinal obstruction group, 2-day natural recovery group, 4-day natural recovery group, 2-day treated group, and 4-day treated group. Sijunzi decoction was given twice a day to the treated groups. The concentration of markers was analyzed to evaluate the immune function and nutritional status. The concentration of interleukin-2, immunoglobulins and complement components of the treated groups were significantly higher than the natural recovery group (P<0.05). The levels of CD4+ and CD4+/CD8+ increased then decreased in the treated groups. The levels of tumor necrosis factor-α and CD8+ were significantly lower than the natural recovery group. The level of total protein in the treated groups also increased then decreased after relieving the obstruction. The levels of albumin, prealbumin and insulin-like growth factor-1 were significantly higher in the treated groups than in the natural recovery group (P<0.05). Transferrin level in the treated groups was significantly higher than the obstruction group (P<0.05). Sijunzi decoction can lessen the inflammatory response and improve the nutrition absorption after relieving the obstruction.

Comparative pathogenicity of Coxsackievirus A16 circulating and noncirculating strains in vitro and in a neonatal mouse model

An enterovirus 71 (EV71) vaccine for the prevention of hand, foot, and mouth disease (HMFD) is available, but it is not known whether the EV71 vaccine cross-protects against Coxsackievirus (CV) infection. Furthermore, although an inactivated circulating CVA16 Changchun 024 (CC024) strain vaccine candidate is effective in newborn mice, the CC024 strain causes severe lesions in muscle and lung tissues. Therefore, an effective CV vaccine with improved pathogenic safety is needed. The aim of this study was to evaluate the in vivo safety and in vitro replication capability of a noncirculating CVA16 SHZH05 strain. The replication capacity of circulating CVA16 strains CC024, CC045, CC090 and CC163 and the noncirculating SHZH05 strain was evaluated by cytopathic effect in different cell lines. The replication capacity and pathogenicity of the CC024 and SHZH05 strains were also evaluated in a neonatal mouse model. Histopathological and viral load analyses demonstrated that the SHZH05 strain had an in vitro replication capacity comparable to the four CC strains. The CC024, but not the SHZH05 strain, became distributed in a variety of tissues and caused severe lesions and mortality in neonatal mice. The differences in replication capacity and in vivo pathogenicity of the CC024 and SHZH05 strains may result from differences in the nucleotide and amino acid sequences of viral functional polyproteins P1, P2 and P3. Our findings suggest that the noncirculating SHZH05 strain may be a safer CV vaccine candidate than the CC024 strain.

The changes of muscle FGF-6 expression at different time points after exercise-induced muscle damage / Cambios de expresión de FGF-6 muscular en diferentes puntos del tiempo después del daño muscular inducido por ejercicio

OBJECTIVE: To investigate the changes of fibroblast growth factor (FGF)-6 expression in the regeneration and repair process after exercise-induced muscle damage (EIMD) and the relationship with skeletal muscle regeneration and repair. METHODS: The expression of FGF-6 at different time points was examined by reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemistry staining after a downhill treadmill exercise. Skeletal muscle injury and regeneration at different times after EIMD was assessed by haematoxylin and eosin (H & E) staining. RESULTS: The FGF-6 protein expression was initially elevated, followed by a gradual reduction, while the changes of FGF-6 mRNA were almost all raised after the treadmill exercise. CONCLUSION: The results point out that FGF-6 is closely related to skeletal muscle regeneration and repair, probably implying a dual function in muscle regeneration.

OBJETIVO: Investigar los cambios de expresión del factor de crecimiento fibroblástico (FGF)-6 en el proceso de regeneración y reparación después de daño muscular inducido por el ejercicio (DMIE) y la relación con la reparación y regeneración del músculo esquelético. MÉTODOS: La expresión de FGF-6 en diferentes tiempos fue examinada mediante reacción en cadena de la polimerasa con transcriptasa inversa (RT-PCR) y tinción inmunohistoquímica, después de un ejercicio de carrera descendente en cinta rodante. La lesión del músculo esquelético y la regeneración en diferentes momentos después del DMIE, fueron evaluadas mediante hematoxilina y eosina (H & E). RESULTADOS: La expresión de la proteína FGF-6 fue elevada al principio, seguida por una reducción gradual, mientras que los cambios de FGF-6 mRNA fueron casi todos incrementados tras los ejercicios en la cinta rodante. CONCLUSIÓN: Nuestros resultados señalan que FGF-6 se relaciona estrechamente con la regeneración del músculo esquelético, lo que probablemente implica una función dual en la regeneración muscular.

Involvement of Src tyrosine kinase and protein kinase C in the expression of macrophage migration inhibitory factor induced by H2O2 in HL-1 mouse cardiac muscle cells

Macrophage migration inhibitory factor (MIF), a pleiotropic cytokine, plays an important role in the pathogenesis of atrial fibrillation; however, the upstream regulation of MIF in atrial myocytes remains unclear. In the present study, we investigated whether and how MIF is regulated in response to the renin-angiotensin system and oxidative stress in atrium myocytes (HL-1 cells). MIF protein and mRNA levels in HL-1 cells were assayed using immunofluorescence, real-time PCR, and Western blot. The result indicated that MIF was expressed in the cytoplasm of HL-1 cells. Hydrogen peroxide (H2O2), but not angiotensin II, stimulated MIF expression in HL-1 cells. H2O2-induced MIF protein and gene levels increased in a dose-dependent manner and were completely abolished in the presence of catalase. H2O2-induced MIF production was completely inhibited by tyrosine kinase inhibitors genistein and PP1, as well as by protein kinase C (PKC) inhibitor GF109203X, suggesting that redox-sensitive MIF production is mediated through tyrosine kinase and PKC-dependent mechanisms in HL-1 cells. These results suggest that MIF is upregulated by HL-1 cells in response to redox stress, probably by the activation of Src and PKC.

Genetic analysis of the ELOVL6 gene polymorphism associated with type 2 diabetes mellitus

Recent animal studies have indicated that overexpression of the elongation of long-chain fatty acids family member 6 (Elovl6) gene can cause insulin resistance and β-cell dysfunction. These are the major factors involved in the development of type 2 diabetes mellitus (T2DM). To identify the relationship between single nucleotide polymorphisms (SNP) of ELOVL6 and T2DM pathogenesis, we conducted a case-control study of 610 Han Chinese individuals (328 newly diagnosed T2DM and 282 healthy subjects). Insulin resistance and islet first-phase secretion function were evaluated by assessment of insulin resistance in a homeostasis model (HOMA-IR) and an arginine stimulation test. Three SNPs of the ELOVL6 gene were genotyped with polymerase chain reaction-restriction fragment length polymorphism, with DNA sequencing used to confirm the results. Only genotypes TT and CT of the ELOVL6 SNP rs12504538 were detected in the samples. Genotype CC was not observed. The T2DM group had a higher frequency of the C allele and the CT genotype than the control group. Subjects with the CT genotype had higher HOMA-IR values than those with the TT genotype. In addition, no statistical significance was observed between the genotype and allele frequencies of the control and T2DM groups for SNPs rs17041272 and rs6824447. The study indicated that the ELOVL6 gene polymorphism rs12504538 is associated with an increased risk of T2DM, because it causes an increase in insulin resistance.

Late onset systemic lupus erythematosus with severe hypercalcaemia / Lupus eritematoso sistémico de inicio tardío con hipercalcemia severa

We report a case of a 76-year old female presenting with symptomatic severe hypercalcaemia, and subsequently diagnosed with late onset SLE due to the presence of anaemia, leucopenia, antibodies of antinuclear (ANA), anti-dsDNA, and also kidney impairment. Serum levels of FGF23 and intact-parathyroid hormone (iPTH) were low in this patient. Serum calcium, FGF23 and iPTH levels responded to steroids, which occurred simultaneously with disease activity. On follow-up, the faster increase in FGF23 than in parathyroid hormone suggested that FGF23 might be involved in the pathogenesis of hypercalcaemia in SLE.

Se reporta el caso de una mujer de 76 años de edad que se presentó con hipercalcemia sintomática severa, y a la que posteriormente le fuera diagnosticada LES de inicio tardío con presencia de anemia, leucopenia, anticuerpos antinucleares (ANA), anti-dsDNA, e insuficiencia del riñón. Los niveles séricos del factor de crecimiento fibroblástico 23 (FGF23) y la hormona paratiroidea intacta (iPTH) fueron bajos en este paciente. Los niveles de calcio séricos, FGF23 e iPTH respondieron a los esteroides, que ocurrieron simultáneamente con la actividad de la enfermedad. En el seguimiento, el hecho de que el factor FGF23 aumentara más rápidamente que la hormona paratiroidea, sugiere que el FGF23 podría estar involucrado en la patogénesis de la hipercalcemia en LES.

Noggin induces human bone marrow-derived mesenchymal stem cells to differentiate into neural and photoreceptor cells.

The present study was undertaken to explore the effect of noggin on neuronal differentiating potential of human bone marrow-derived mesenchymal stem cells (hBMMSCs) in vitro so as to provide a means of alleviate retinal degeneration. A green fluorescent protein-tagged noggin gene was transferred into adult hBMMSCs or induce hBMMSCs with classical inducer, epidermal growth factor(EGF). Neurons were observed as early as 48 h after transduction of hBMMSCs with a noggin adenoviral vector. Differentiation peaked by 10 days in culture, and these differentiated cells expressed multiple markers including rhodopsin (18.4 ± 1.5% of cells), chx10 (4.8 ± 0.6%), nestin (4.2 ± 0.8%), and Nrl (3.7 ± 0.4%), as verified by immunofluorescence staining. Noggin-transduced cells produced more photoreceptor cells than non-transduced cells, suggesting that noggin has the ability to induce hBMMSCs to trans-differentiate into photoreceptor cells. In contrast, induction with EGF for 10 days led to lower levels of rhodopsin and chx10, and undetectable levels of Nrl and Nestin. These findings suggested noggin-transduced hBMMSCs produced more photoreceptor cells than EGF–induced cells. It is suggested that the present protocol has application in cell replacement therapy for patients suffering from photoreceptor cell loss.

Proteomic analysis of clear cell renal cell carcinoma. identification of potential tumor markers

To identify new tumor specific proteins of renal cell carcinoma [RCC] using proteomic analysis. Nine renal cell carcinomas were resected and these surgical operations were carried out from June 2007 to September 2008 in the Urology Department of the Second Affiliated Hospital of Soochow University, Suzhou, Jiangsu Province, China. The cancer tissues and para-cancer normal tissues were preserved in liquid nitrogen. We analyzed the RCC tissues and para-cancer normal tissues by 2-dimensional polyacrylamide gel electrophoresis [2D-PAGE]. The 16 differentially expressed protein spots [p<0.05 by Student t-test] were identified by matrix assisted laser desorption ionization/time of flight mass spectrometry and tandem mass spectrometry. Six proteins were down regulated and 10 proteins were up regulated in clear cell RCC compared with corresponding normal kidney tissue. The down regulated proteins were calbindin, ester hydrolase C11orf54, alcohol dehydrogenase, ammecr1-like protein, adenosine diphosphate [ADP]/adenosine-5'-triphosphate [ATP] translocase 3 and leucine-tyrosine-arginine [LYR] motif-containing protein 5. The up regulated proteins were hypoxia inducible domain family member 1A, glutathione S-transferase P, thioredoxin-dependent peroxide reductase, peroxiredoxin-6, CD2-associated protein, annexin A5, gamma-enolase, retinal dehydrogenase 1, vimentin, and protein-glutamine gamma-glutamyltransferase 2. The data provide potential tumor markers for diagnosis of RCC

Field assessment of recombinant Schistosoma japonicum 26 kDa glutathione S-transferase in Chinese water buffaloes.

We have shown previously that anti-fecundity immunity can be induced experimentally against recombinant 26 kDa glutathione S-transferase (reSjc26GST) in Chinese water buffaloes (Bos buffelus), important reservoir hosts for Schistosoma japonicum in China. In the field study described here, we immunized buffaloes with reSjc26GST to induce protective immunity against S. japonicum and to evaluate its effectiveness in controlling schistosomiasis japonica. We selected two villages as test and control groups in inside-embankment areas endemic for schistosomiasis japonica. The buffaloes in the test village were vaccinated with reSjc26GST, whereas those in the control village were not. The indicators of the effect of the vaccine included the generation of specific IgG antibodies in the vaccinated buffaloes, changes in the prevalence and infection intensity in buffaloes and village children, changes in the density of infected snails, and changes in the infectivity of water bodies (assessed by sentinel mice) in transmission areas adjacent to both villages. Twenty months after vaccination, the infection rate of buffaloes in the test village was decreased by 60.4% (from an initial prevalence of 13.5% to 5.4%), and 67.9% when compared with that in the control village (initial prevalence of 16.7%). However, the infection rate in village children remained unchanged. The density of infected snails decreased by 71.4%, from 0.0049/0.11 m2 to 0.0014/0.11m2 in the high transmission area outside the embankment in the test village. There was no change in the infectivity of the water body transmission areas between the test and control villages. The levels of specific antibodies to reSjc26GST showed a continuous increase after vaccination. These results indicate that protective immunity was induced and maintained in buffaloes after vaccination with reSjc26GST. The vaccine could thus play a significant role in reducing S. japonicum transmission caused by water buffaloes in the Lake region of China.

Causes et traitement des complications des suites operatoires de l'oeil glaucomateux

Sur les 43 glaucomes (28 malades) ayant subi une operation de filtration cites dans cet article; 12 yeux ont la complication de chambre anterieure superficielle. Cette complication est causee principalement par le diastasis choroido-ciliaire et l'exces de filtration. Avec le moyen de traitement presente ci-apres; la chambre anterieure sera retablie a 100 pour cent. La chambre anterieure superficielle apres filtration du glaucome est une complication immediate et courante apres l'operation. Si elle n'est pas bien traitee; elle va faire mauvais effet a l'operation du glaucome. Pour faire des recherches sur cette complication; on a fait des analyses sur les glaucomes traites depuis 1994